Copyright
National Institute of Environmental Health Science.

Annual report : National Institute of Environmental Health Sciences (Volume 1982) online

. (page 21 of 90)
Online LibraryNational Institute of Environmental Health ScienceAnnual report : National Institute of Environmental Health Sciences (Volume 1982) → online text (page 21 of 90)
Font size
QR-code for this ebook


makes a demonstration of its existence in a biological system of considerable
importance.



236



SMITHSONIAN SCIENCE INFORMATION EXCHANGE
PROJECT NUMBER (Oo NOT use this space)



U.S. DEPARTMENT OF

HEALTH AND HUMAN SERVICES

PUBLIC HEALTH SERVICE

NOTICE OF

IHTRAHURAL RESEARCH PROJECT



PROJECT NUMBER



ZOl ES 50056-03 LEB



PERIOD COVERED

O£tober 1,2.981 to^ September 30, 1982_

TITLE OF PROJECT (eo'charactcrs or loss)

Effects of hypothermia on ion movement in guinea pig cochlea



NAMES, LABORATORY AND INSTITUTE AFFILIATIONS, AND TITLES OF PRINCIPAL INVESTIGATORS AND ALL OTHER
PROFESSIONAL PERSONNEL ENGAGED ON THE PROJECT



PI:



OTHER:



Teruzo Konishi
Alec N. Salt



LEB
LEB



Philip E. Hamrick SFTY



Medical Officer (Research)
Visiting Fellow

Radiation Safety Officer



COOPERATING UNITS (if any)

Radiation Safety



lab/branch

Laboratory of Environmental Biophysics



SECTION

Noise Effects Research Workgroup



INSTITUTE AND LOCATION

NIEHS. NIH. Research Triangle Park, N.C. 27709



TOTAL MANYEARS:





PROFESSIONAL:





CHECK APPROPRIATE BOX(ES)
D (a) HUMAN SUBJECTS

D (al) MINORS n (a2) INTERVIEWS



□ (b) HUMAN TISSUES



(c) NEITHER



SUMMARY OF WORK (200 words or less - underline keywords)

It has been documented that the cochlear potentials are maintained by metabolic
energy. However, it is not known whether a decrease of metabolic rate by hypo-
thermia can alter ion movement or membrane permeability of the cochlea . The
present study is designed to determine dependence of cochlear membrane perme-
ability on temperature.



237



PHS-6040
(Rev. 2-81)



Z01 ES 50056-03 LEB

diffusion. The K conductance was calculated from the rate of change of the endo-
lymph K concentration relative to the K electrochemical potential difference.
The mean G,, averaged from 10 to 30 min after onset of anoxia was (21.53 + 5.54)x
10"6mho in hypothermic guinea pigs, whereas the mean G|. averaged during the same
period was (34.85 +; 5.60)xl0 ^mho in normal guinea pigs.

A manuscript has been accepted for publication and this project has been completed.

SIGNIFICANCE TO BIOMEDICAL RESEARCH AND THE PROGRAM OF THE INSTITUTE : The present
data suggest that hypothermia results in changes in both active and passive ion
transport mechanisms in the cochlea. It is likely that a decrease of metabolic
energy caused by ototoxic insults may interfere not only with active ion transport,
but also the membrane permeability of the cochlear partition.

PUBLICATIONS

Komshi, T., Salt, A.N. and Hamrick, P.E.: Effects of hypothermia on ion movement
in guinea pig cochlea. Hearing Res. 4^: 265-278, 1981.



238



Z01 ES 50056-03 LEB

PROJECT DESCRIPTION

METHODS EMPLOYED : The cochlear potentials were recorded from the basal turn of
the cochlea of guinea pigs anesthetized with sodium pentobarbital. The rectal
temperature and blood pressure in the common carotid artery were monitored. The
rectal temperature was reduced from 39° + 0.5°C to 29° + 0.5°C with a cooling
pad. The perfusion of the perilymphatic space with solutions containing ^'^K and
^^Na and collection of the cochlear fluids were carried out during steady state
hypothermia. Total concentrations of K and Na in the cochlear fluids were
determined by a helium glow photometer and radioactivities of "^^k and 22|\ja ^^ere
determined by gamma spectrometry.

MAJOR FINDINGS AND PROPOSED COURSE : 1. Cochlear potenitals. The endocochlear
potential (EP) measured at 39°C was 86.0 + 3.5 mV. When the rectal temperature
decreased to 29°C the EP was 78.2 + 3.5 mV. The EP recorded 2 hours after the
rectal temperature was kept at 29.0° + 0.5°C was 73.2 + 3.9 mV. During the course
of hypothermia the magnitude of cochlear microphonics TCM) gradually decreased.
Usually a large increase of the negative summating potential accompanied the
decrease in CM during the period of hypothermia. The action potential (AP) in
response to test stimuli of low intensity were markedly suppressed but the AP
remained little changed or became supernormal with high intensity stimuli.

2. Electrolyte concentrations in the cochlear fluids. Hypothermia did not
result in marked changes in K concentrations in the endloymph and perilymph of
nonperfused cochlea. The Na concentrations in both endolymph and perilymph were
slightly decreased in hypothermic guinea pigs.

3. Membrane permeability of the endolymph-perilymph barrier determined by rate
constant for K. When the perilymphatic space was perfused with radioactive
artificial perilymph, the ^^k concentrations in the endolymph (normalized by ^'^K
concentrations in_the perilymph) increased exponentially. The rate constant for
K was 0.0069 min"^ in hypothermic guinea pigs which was significantly lower than
the value obtained in normal guinea pigs (0.013 min ^).

The K conductance of the endolymph-perilymph barrier can be computed by using
the following equation

„ K end "- end""



Ay



where V , is volume of the endolymph (2m1) and Ay' is the electrochemical potential
difference between endolymph and perilymph. The computed G,, was 20.65 x 10"^
mho in hypethermic guinea pigs which was considerably lower than 35.16 x 10"^ mho
in normal guinea pigs. The permeability constant for K (P^^) was 133.54 x 10"^
cm3 sec"i in hypothermic guinea pigs. This value was substantially lower than
the normal value (242.9 x lO""* cm^ sec M-

4. Membrane permeability of the endolymph-perilymph barrier determined from k"*"
passive diffusion during anoxic period. Permanent anoxia was induced 30 min
after the rectal temperature fell to 29.0"^. When anoxia continued for longer
than 5 min the decrease of the endolymph K concentration measured with K
selective liquid membrane electrodes was solely attributable to a passive K^

239



MITHSONIAW SCIENCE INFORMATION EXCHANGE
PROJECT NUMBER (Oo NOT use ihis space)



U.3. DEPARTMENT OF

HEALTH AND HUMAN SERVICES

PUBLIC HEALTH SERVICE

NOTICE OF

INTRAMURAL RESEARCH PROJECT



PERIOD COVERED

Octob er 1, 19 81 to September 30, 1982

TITLE OF PROJECT Ilo characters or less)

Effects of Microwave Radiation on Reproductive Cells



PROJECT NUMBER



ZOl ES 50057-03 LEB



NAMES, LABORATORY AND INSTITUTE AFFILIATIONS, AND TITLES OF PRINCIPAL INVESTIGATORS AND ALL OTHER
PROFESSIONAL PERSONNEL ENGAGED ON THE PROJECT

PI: Michael J. Galvin Senior Staff Fellow LEB NIEHS



OTHERS:



Donald I. McRee
Cindy H. Hall
J. Paul Thaxton



Research Physicist
Graduate Student
Poultry Science Dept.



LEB NIEHS

N.C. State University

N.C. State University



COOPERATING UNITS (if any)



None



lab/branch

Laboratory of Environmental Biophysics



SECTION

Non-Ionizing Radiation Workgroup



INSTITUTE AND LOCATION

NIEHS. NIH, Research Triangle. N.C,



TOTAL MANYEARS:

0.2



PROFESSIONAL:

0.1



CHECK APPROPRIATE 80X(ES)
D (a) HUMAN SUBJECTS

D (al) MINORS n (a2) INTERVIEWS



OTHER:



0.1



D (b) HUMAN TISSUES



)D (c) NEITHER



SUMMARY OF WORK (200 words or less - underline keywords)

In this study, the effect of non- ionizing radiation on the integrity of mature
spermatocytes was examined. Semen was obtained from 10-month-old turkeys and
diluted to a concentration of 3.5 x 10^ sperm/ml using Beltsville Poultry Semen
Extender. The sperm were exposed for 30 minutes to 2.45 GHz microwave radiation
at specific absorption rates (SAP's) of 0, (sham) 10 or 50 mW/g. . The waveguide
exposure system maintained the control (non-irradiated) and irradiated samples at
40 + 0.1 5°C throughout the experiment. Microwave radiation did not alter membrane
permeability to a vital stain or the intracellular enzymes LDH and GOT. The iji
vivo performance of the sperm following in vitro irradiation was also examined.



The fertility of virgin turkey hens given a single insemination of microwave
exposed sperm was similar for the 10 mW/g and nonexposed sperm. Hatchability
and embryonic mortality were not different among the treatment groups. A number
of hematological parameters in the progeny of the turkey hens are now being
examined.



240



PHS-6040
(Rev. 2-81)



Z01 ES 50057-03 LEB

PROJECT DESCRIPTION

METHODS EMPLOYED : a. Cell Preparation : Semen was obtained from 7-month-old
Nicholas large white turkeys. Biological variation was minimized by pooling se-
men from 30 turkeys. For each experiment, 2 ml of semen were collected and
diluted 2:1 with Beltsville Poultry Semen Extender, BPSE (USDA, Beltsville, MD).
The semen was washed twice to remove the seminal plasma. The sperm were then
resuspended in BPSE to a concentration of 5.0 x 10^ sperm/ml. The sperm were
irradiated in an S-band waveguide chamber at a frequency of 2450 MHz. Each tube
contained 4.2 ml of the sperm suspension. The tubes were siliconized to mini-
mize cellular adhesion and were gently stirred throughout the exposure duration
to allow uniform microwave absorption and temperature distribution.

b. Cellular Integrity : Several parameters were selected as indices of altered
membrane function: permeability to a vital stain (viability) release of lactate
dehydrogenase (LDH) and release of glutamic oxalic transaminase (GOT).

c. In vivo performance of sperm following Jjl vjtro irradiation: Virgin turkey
hens were given a single insemination of sperm which were either sham-exposed, or
exposed to 10 mW/g or 50 mW/g. Egg fertility was determined during the nine
weeks following insemination.

d. Evaluation of progeny from turkey hens receiving microwave irradiated sperm.
Hematological evaluation of the progeny from eggs laid during the second and
fourtli weeks after insemination. The turkey polts were evaluated a 2, 4 and

5 weeks of age.

MAJOR FINDINGS AND PROPOSED COURSE : a. Cellular Integrity : Initially, the
viability was 96% and was not affected by microwave radiation at any of the
exposure levels examined. The release of the soluble enzymes, lactic acid
dehydrogenase, and glutamic oxalic transaminase, by sperm into the suspending
media was not altered.

b. In vivo performance of sperm following in vitro irradiation : Fertility de-
clined comparably in all groups until the fourth week after insemination. During
the fourth week, fertility increased in the hens which received 50 mW exposed sperm
and elevated fertility was maintained in this group for the duration of the
experiment. Hatchability and embryonic mortality were not different.

c. Evaluation of progeny from turkey hens receiving microwave irradiated sperm :
No data available.

SIGNIFICANCE TO BIOMEDICAL RESEARCH AND THE PROGRAM OF THE INSTITUTE : The
potential health effects of microwave radiation in the environment are of inter-
est to the National Institute of Environmental Heolth Sciences. Before an
accurate evaluation of the biological effects of 2450 MHz microwaves can be
made, it is necessary to control the temperature of the specimen carefully and
be able to reproduce the exposure conditions. We have developed the capability
to do this for in vitro exposures at NIEHS and this may provide a system for
differentiating specific microwave effects from thermal responses. In addition,
by examining the response of cells and cellular components to microwave radiation,

24!



ZOl ES 50057-03 LEB



it should be easier to identify the mechanisms of action of microwave radiation
with biological specimens.



242



SMITHSONIAN SCIENCE INFORMATION EXCHANGt U.S. DEPARTMENT OF

PROJECT NUMBER (Do NOT use this space) 1] HEALTH AND HUMAN SERVICES

i PUBLIC HEALTH SERVICE

I NOTICE OF

I INTRAHURAL RESEARCH PROJECT



PROJECT NUMBER



ZOl ES 50058-03 LEB



PERIOD COVERED

Oc to ber 1, 1981 to September 30. 1982



TITLE OF PROJECT (80 characters or less)

Microwave Effects on Fetal Development in Mice



NAMES, LABORATORY AND INSTITUTE AFFILIATIONS, AND TITLES OF PRINCIPAL INVESTIGATORS AND ALL OTHER
PROFESSIONAL PERSONNEL ENGAGED ON THE PROJECT



PI



OTHER;



Donald I . McRee
Peter Nawrot

Minuro Inouye
R.E. Staples



Research Physicist
Visiting Associate

Visiting Fellovj
Research Scientist



LEB
LEB



NIEHS
NIEHS



LEB NIEHS

Dupont Corp. ,
Wilmington, Del .



COOPERATING UNITS (if any)

Research Triangle Institute



lab/branch

Laboratory of Environmental Biophysics



SECTION

Non-Ionizing Radiation Workgroup



INSTITUTE AND LOCATION

NIEHS. NIH, Research Triangle Park, N.C. 27709



TOTAL MANYEARS:

0.7



PROFESSIONAL:

0.5



OTHER:



0.2



CHECK APPROPRIATE BOX(ES)

□ (a) HUMAN SUBJECTS

□ (al) MINORS D (a2) INTERVIEWS



□ (b) HUMAN TISSUES



H (c) NEITHER



SUMMARY OF WORK (200 words or less - underline keywords)

Pregnant mice (CD-I strain) were exposed to 2.45 GHz microwave radiation at a
power density level of 30 mW/cm^. At exposure to 30 mW/cm^ (SAR : 32 mW/g)
during days 1-6 a significant decrease in implantation sites per litter and
average fetal weight was observed. Exposure to 30 mW/cm^ during days 6-15
resulted in a slight increase in the number of malformed fetuses but was not
statistically significant as obtained in a previous experiment. This repeat
of previous work again indicates that the threshold for teratogenic effects in
the CD-I mouse strain is approximately 30 mW/cm^.



243



PHS-6040
(Rev. 2-81)



ZOl ES 50058-03 LEB



PROJECT DESCRIPTION

METHODS EMPLOYED : The objective of this research was to determine the maternal
and embryotoxic effects of microwaves. In order to determine whether or not the
effects were only thermal or a combination of thermal and specific microwave
interactions, groups of animals were placed in elevated temperature environments
in order to simulate the thermal stress of the microwave exposure. The mice were
exposed from above in styrofoam cages (one animal per cage) separated at least 2
body lengths with the long axis of the cages parallel to the electric field.

MAJOR FINDINGS AND PROPOSED COURSE : We repeated the experiment which exposed
pregnant CD-I mice to 30 mW/cm^ to 2.45 - GHz microwave radiation. Our investi-
gation showed that exposure to 30 mW/cm^ during days 1-6 produced a signficant
decrease in implantation sites per litter and average fetal weight. In our
previous study exposure to 30 mW/cm^ during days 6-15 resulted in a small but
statistically significant increase in the number of malformed fetuses, primarily
cleft palate. In this study a slight but not significant increase in malformed
fetuses were observed. This supports our original contention that 30 mW/cm^ (SAR
; 32 mW/g) is near the threshold level for producing teratogenic effects in CD-I
mice.

The research project will be completed during this fiscal year.

SIGNIFICANCE TO BIOMEDICAL RESEARCH AND THE PROGRAM AT THE INSTITUTE: Sufficient
information is not available at the present time to establish scientifically
based safety standards for microwave radiation exposure. The determination of
the intensity of microwave radiation which produces teratogenic effects is
important to the evaluation of the hazardous effects of microwaves. This research
is part of the mission of the Institute to conduct research on the health effects
of physical factors in the environment.

PUBLICATIONS

Nawrot, P.S., McRee, D.I. and Staples, R.E.: Effects of 2.45 GHz microwave
radiation on embrvofetal development in mice. Teratology 24: 303-314, 1981.



244



SMITHSONIAN SCIENCE INFORMATION EXCHANGE
PROJECT NUMBER (Do NOT use this space)



PERIOD COVERED

October 1, 1981 to September 30, 1982



U.S. DEPARTMENT OF

HEALTH AND HUMAN SERVICES

PUBLIC HEALTH SERVICE

NOTICE OF

INTRAMURAL RESEARCH PROJECT



PROJECT NUMBER



ZOl ES 50059-03 LEB



TITLE OF PROJECT (80 characters or less)

Protoporphyrin Phototoxicity in Rat Mast Cells and Human Erythrocytes



NAMES, LABORATORY AND INSTITUTE AFFILIATIONS, AND TITLES OF PRINCIPAL INVESTIGATORS AND ALL OTHER
PROFESSIONAL PERSONNEL ENGAGED ON THE PROJECT



PI : Mary J. Ortner
OTHER: Colin F. Chignell



Senior Staff Fellow


LEB


NIEHS


Chief


LEB


NIEHS



COOPERATING UNITS (if any)



Mone



las/branch

Laboratory of Environmental Biophysics



ECTION

Molecular Biophysics



INSTITUTE AND LOCATION

NIEHS, NIH, Research Triangle Park, North Carolina 27709



TOTAL MANYEARS:

0.7



PROFESSIONAL:

0.2



0.5



check APPROPRIATE BOX(ES)
D (a) HUMAN SUBJECTS

□ (al) MINORS D (a2) INTERVIEWS



n (b) HUMAN TISSUES



n (c) NEITHER



I



SUMMARY OF WORK (200 words or less - underline keywords)

Pro toporphyr in-mediated phototoxicity has been studied in human erythrocytes and
rat mast ceil s. Cir'cuTa'r dichroism studies of erythrocyte ghosts have shown that
phototox"ic lysis is accompanied by membrane protein denaturation with loss of
alpha helical structure. Rat mast cells showed a dual phototoxic response which
depended on the light intensity. Low intensity light caused a stabilization
of the cell membrane resulting in a loss of histamine secretory ability;
whereas high intensity light caused phototoxic lysis . Sodium dodecyl sulfate
disc gel electrophoresis indicated that the mast cell membrane proteins are ,
covalently crossl inked by the phototoxic reaction in a manner similar to that
seen in erythrocyte ghosts. Biophysical studies are currently underway to
establish the molecular mechanisms of these phenomonea.



245



PHS-6040
(Rev. 2-81)



ZOl ES 50059-03 LEB



PROJECT DESCRIPTION



OBJECTIVES : Severe phototoxic reactions may be clinically manifested in patients
with erythropoietic porphoryia, a metabolic disease which results in a high
buildup of porphyrins in the blood. Porphyrin-mediated phototoxicity is related
to erythrocyte hemolysis, and in spite of extensive biochemical studies, the
molecular mechanism is not completely understood. We have studied this photo-
toxic reaction in erythrocyte ghosts using circular dichroism (CD), a technique
which is sensitive to membrane protein conformation. In addition, we have
studied the reaction in a eukaryotic cell which can be stimulated in vitro to
perform a biological response. The purpose of this approach is to study more
closely the intermediate oxidative reactions which precede erythrocyte lysis, and
to determine the effects of these reactions on erythrocytes and functioning
eukaryotic cells.

METHODS EMPLOYED : Purified rat peritoneal mast cells and human erythrocyte
ghosts were obtained using well established methods. Cells and ghosts were
exposed to light in the presence of protoporphyrin using either a lOOW incandescent
light bulb or a lOOW mercury vapor lamp. The light intensity was varied by
changing the distance from the light source. Circular dichroism measurements
were performed on erythrocyte ghosts exposed to light and protoporphyrin using a
Jasco automatic recording polarimeter. Simultaneous measurements of membrane
protein optical density at 280nm indicated that changes in the circular dichroism
spectra were not due to scattering artifacts.

MAJOR FINDINGS : Human erythrocytes and ghosts. Human erythrocytes exposed to
strong light in the presence of protoporphyrin underwent lysis on a time-dependent
basis. An equivalent number of ghosts exposed under identical conditions showed
changes in CD spectra of the membrane proteins which were consistent with loss in
a-helical structure due to protein denaturation. Appropriate controls showed
that the effect was dependent both on protoporphyrin and light. Changes in the
CD spectra began after 2-4 min when the ghosts (20 yg Protein/ml) were exposed to
2mM protoporphyrin and placed 20 cm from the mercury vapor light source. In the
intact erythrocytes, phototoxic lysis began between 6-8 min after the begining of
illumination and progressed with time in a sigmoidal fashion until almost 100%
lysis occurred. The data therefore indicate that changes in the secondary protein
structure of erythrocyte ghost proteins can be correlated u'ith protoporphyrin
induced photolysis.

Although D2O decreased the rate of lysis in erythrocyte ghosts, there were no
effects on the rate of decrease in membrane protein- secondary structure as deter-
mined by circular dichroism. This suggests that either singlet oxygen does not
play a significant role in protoporphyrin phototoxicity or that in this system,
singlet oxygen does not decay via collisions with the aqueous environemnt.

Rat Peritoneal Mast Cells . Rat mast cells undergo lysis when exposed to proto-
porphyrin and high intensity light. This reaction is manifested by the cytotoxic
release of histamine due to conditions which may be similar to those seen in
erythrocyte ghosts. This study has shown, however, that mast cell membranes do
not lyse when exposed to protoporphyrin and low intensity light, but rather are
stabilized in a way which makes them resistant to histamine liberators. The

246



ZOl ES 50059-03 LEB

development of this inhibition is both dose and time dependent (100 yg/ml proto-
porphyrin for 30 min gives total inhibition) and does not occur in the dark or
under ordinary room light. Purified mast cells were exposed to conditions which
produced inhibition, and the proteins separated using SDS-disc gel electrophoresis,
Preliminary results indicated that some of the proteins were unable to enter the
gel, presumably due to crossl inking and aggregation. This would indicate that
the stabilization phase of protoporphyrin phototoxicity in mast cells may involve
a protein crossl inking similar to that which accompanies erythrocyte lysis.

SIGNIFICANCE TO BIOMEDICAL RESEARCH AND THE PROGRAM OF THE INSTITUTE: Severe
phototoxic urticaria can occur as a result of clinical disorders such as erythro-
poietic porphyria, from direct phototoxic reactions to drugs and chemicals or
from photoallergic reactions to haptens. A biophysical study of phototoxicity in
erythrocyte membranes has not been previously undertaken. Protoporphyrin-induced
phototoxicity is therefore being studied as a model system in order to understand
more clearly the role of active oxygen in the phototoxic response. The use of
these techniques will contribute to our knowledge of both membrane physiology and
phototoxicity. The mast cell provides an excellent model for both in vitro
and in vivo studies of phototoxicity in a eukaryotic cell. Furthermore, mast
cells secrete several mediators of inflammation and since they occur abundantly
in the skin, they may play a direct role in the phototoxic response. A thorough
study is therefore needed to understand the molecular mechanism of this response
in the mast cell. These studies may lead to a method of control over the symptoms
of phototoxic urticaria.

PROPOSED COURSE : We are currently developing techniques to study the effects of
several agents on the early development of membrane protein denaturation and the
involvement of specific ghost proteins. Since protoporphyrin phototoxicity is
probably mediated via reduced forms of active oxygen, we have studied the early
stages of phototoxicity in the presence of agents which are known to affect these
forms (D2O superoxide dismutase, mannitol, catalase, etc.) The use of circular
dichrosim makes it possible to study the time course of membrane protein denatura-
tion directly and also to study this reaction in the early, pre-lytic stages of
phototoxic damage.

The effects of protoporphyrin phototoxicity on membrane intercalated spin labels
will be studied using ESR. Lipid spin labels can be intercalated into erythro-
cyte ghosts to probe the interface regions and the hydrophobic interior. In
addition, the membrane proteins may be covalently spin labeled and the phototoxic
reaction studied at this level. Because the spin probes are degraded by the
strong oxidants produced during the phototoxic response, it may be possible to
determine the area in the membrane where the greatest number of oxidative reac-
tions are occurring, and whether this area changes during the course of the
reaction.

Protoporphyrin mediated phototoxicity has not as yet been studied directly in
erythrocyte ghosts. Using the biophysical techniques outlined above, we hope to
learn more about this phototoxic response and the secondary reactions, it pre-
cipitates in the erythrocyte membrane.

We plan to continue these studies of phototoxicity in mast cells by studying the

247



ZOl ES 50059-03 LEB

reaction in the presence of inhibitors and quenchers of reduced forms of active
oxygen. Although histamine release is affected by some of these agents, the
effects on protein crosslinking may still be determined. In this way, we may
learn whether the phototoxic effect of protoporphyrin is mediated directly or vial



Online LibraryNational Institute of Environmental Health ScienceAnnual report : National Institute of Environmental Health Sciences (Volume 1982) → online text (page 21 of 90)